Ge Liang, Melville David, Zhang Min, Schekman Randy
Department of Molecular and Cell Biology , Howard Hughes Medical Institute, University of California, Berkeley , Berkeley , United States.
Elife. 2013 Aug 6;2:e00947. doi: 10.7554/eLife.00947.
Autophagy is a catabolic process for bulk degradation of cytosolic materials mediated by double-membraned autophagosomes. The membrane determinant to initiate the formation of autophagosomes remains elusive. Here, we establish a cell-free assay based on LC3 lipidation to define the organelle membrane supporting early autophagosome formation. In vitro LC3 lipidation requires energy and is subject to regulation by the pathways modulating autophagy in vivo. We developed a systematic membrane isolation scheme to identify the endoplasmic reticulum-Golgi intermediate compartment (ERGIC) as a primary membrane source both necessary and sufficient to trigger LC3 lipidation in vitro. Functional studies demonstrate that the ERGIC is required for autophagosome biogenesis in vivo. Moreover, we find that the ERGIC acts by recruiting the early autophagosome marker ATG14, a critical step for the generation of preautophagosomal membranes. DOI:http://dx.doi.org/10.7554/eLife.00947.001.
自噬是一种由双膜自噬体介导的对胞质物质进行大量降解的分解代谢过程。启动自噬体形成的膜决定因素仍不清楚。在此，我们建立了一种基于LC3脂化的无细胞分析方法，以确定支持早期自噬体形成的细胞器膜。体外LC3脂化需要能量，并受体内调节自噬的途径调控。我们开发了一种系统的膜分离方案，以确定内质网-高尔基体中间区室（ERGIC）是体外触发LC3脂化既必要又充分的主要膜来源。功能研究表明，ERGIC是体内自噬体生物发生所必需的。此外，我们发现ERGIC通过招募早期自噬体标记物ATG14发挥作用，这是自噬前体膜生成的关键步骤。DOI:http://dx.doi.org/10.7554/eLife.00947.001 。
